This research explores the limitations set by the local microenvironment in vivo on conventional photosensitizers that act via generation of toxic singlet oxygen. Hypoxia is a limiting factor to photodynamic therapy (PDT). Systematic determination of oxygen dependence of the photosensitizers will facilitate better understanding of in vivo photosensitization. Development of alternative, oxygen-independent photodamage sensitization mechanisms that overcome existing limitations in PDT is anticipated. The direct effects of the biological ri~iicroenvironment of the photosensitizer on its mechanism and efficiency will be determined, as well as i the effects of high local sensitizer concentration; direct reaction of photosensitizer intermediates with integral biomolecules and cellular localization sites on the photophysics and phototoxicity of photo-sensitizers will be determined in liposomes, erythrocyte ghosts and malignant cell suspensions. A general relationship will be constructed to predict efficacy of singlet oxygen photo sensitizers to cause cell damage. Diffuse reflectance laser flash photolysis will measure oxygen concentrations in vivo using oxygen-dependent triplet state kinetics. Measured 02 is combined with rate constants for natural decay and oxygen quenching of the triplet state to predict photosensitizer efficiency. This relationship will be tested using photosensitizers with different natural triplet lifetimes, varying 02 through N2 /02 gas mixtures and correlating observed phototoxicity with the efficiency predicted by the photophysical method. A third goal is to develop alternative oxygen-independent photo-sensitization mechanisms for photosensitized damage in biological media that overcome the limitations of hypoxia. Radical species, rather than singlet oxygen, will be generated through bond cleavage and photosensitization processes in related compounds. A correlation of reaction mechanisms with photoio'nization of normal and hypoxic cellular systems will ascertain the potential of radical-based strategies to effect oxygen-independent photodamage.